Protein Adsorption Measurements Research Articles

In Situ Measurement of Nanoparticle-Blood Protein Adsorption and Its Heterogeneity with Single-Nanoparticle Resolution via Dual Fluorescence Quantification.

The formation of a protein corona gives nanomedicines a distinct biological identity, profoundly influencing their fate in the body. Nonspecific nanoparticle-protein interactions are typically highly heterogeneous, which can lead to unique biological behaviors and in vivo fates for individual nanoparticles that remain underexplored. To address this, we have established an in situ approach that allows quantitative examination of nanoparticle-protein adsorption at the individual nanoparticle level. This method integrates dual fluorescence quantification techniques, wherein the nanoparticles are first individually analyzed via nanoflow cytometry to detect fluorescent signals from adsorbed proteins. The obtained fluorescence intensity is then translated into protein quantities through calibration with microplate reader quantification. Consequently, this approach enables analysis of interparticle heterogeneity of nano-protein interactions, as well as in situ monitoring of protein adsorption kinetics and nanoparticle aggregation status in blood serum, preconditioning for a comprehensive understanding of nano-bio interactions, and predicting in vivo fate of nanomedicines.

Competitive protein adsorption in isocratic anion exchange chromatography

Protein chromatography is the dominant method of purification of biopharmaceuticals. Although all practical chromatography involves competitive absorption and separation of M. species, competitive protein absorption has remained inadequately understood. We previously introduced the measurement of equilibrium protein adsorption isotherms with all intensive variables held constant, including competitor concentration. In this work, we introduce isocratic chromatographic retention measurements of dynamic protein adsorption in the presence of a constant concentration of a competitor protein. These measurements are achieved by establishing a dynamic equilibrium with a constant concentration of competitor (insulin) in the mobile phase flowing through an ion exchange adsorbent column and following the behavior of a test protein (α-lactalbumin) injected into this environment. We observed decreased retention times for α-lactalbumin in presence of the competitor. The presence of competitor also reduces the heterogeneity of the sites available for adsorption of the test protein. This investigation provides an approach to fundamental understanding of competitive dynamics of multicomponent protein chromatography.

Co-Electrospun Poly(ε-Caprolactone)/Zein Articular Cartilage Scaffolds.

Osteoarthritis scaffold-based grafts fail because of poor integration with the surrounding soft tissue and inadequate tribological properties. To circumvent this, we propose electrospun poly(ε-caprolactone)/zein-based scaffolds owing to their biomimetic capabilities. The scaffold surfaces were characterized using Fourier-transform infrared spectroscopy, X-ray photoelectron spectroscopy, static water contact angles, and profilometry. Scaffold biocompatibility properties were assessed by measuring protein adsorption (Bicinchoninic Acid Assay), cell spreading (stained F-actin), and metabolic activity (PrestoBlue™ Cell Viability Reagent) of primary bovine chondrocytes. The data show that zein surface segregation in the membranes not only completely changed the hydrophobic behavior of the materials, but also increased the cell yield and metabolic activity on the scaffolds. The surface segregation is verified by the infrared peak at 1658 cm-1, along with the presence and increase in N1 content in the survey XPS. This observation could explain the decrease in the water contact angles from 125° to approximately 60° in zein-comprised materials and the decrease in the protein adsorption of both bovine serum albumin and synovial fluid by half. Surface nano roughness in the PCL/zein samples additionally benefited the radial spreading of bovine chondrocytes. This study showed that co-electrospun PCL/zein scaffolds have promising surface and biocompatibility properties for use in articular-tissue-engineering applications.

Responses of Rat Mesenchymal Stromal Cells to Nanocellulose with Different Functional Groups.

Cellulose nanofibrils (CNFs) are multiscale hydrophilic biocompatible polysaccharide materials derived from wood and plants. TEMPO-mediated oxidation of CNFs (TO-CNF) turns some of the primary hydroxyl groups to carboxylate and aldehyde groups. Unlike carboxylic functional groups, there is little or no information about the biological role of the aldehyde groups on the surface of wood-based CNFs. In this work, we replaced the aldehyde groups in the TO-CNF samples with carboxyl groups by another oxidation treatment (TO-O-CNF) or with primary alcohols with terminal hydroxyl groups by a reduction reaction (TO-R-CNF). Rat mesenchymal stem/stromal cells (MSCs) derived from bone marrow were seeded on polystyrene tissue culture plates (TCP) coated with CNFs with and without aldehyde groups. TCP and TCP coated with bacterial nanocellulose (BNC) were used as control groups. Protein adsorption measurements demonstrated that more proteins were adsorbed from cell culture media on all CNF surfaces compared to BNC. Live/dead and lactate dehydrogenase assays confirmed that all nanocellulose biomaterials supported excellent cell viability. Interestingly, TO-R-CNF samples, which have no aldehyde groups, showed better cell spreading than BNC and comparable results to TCP. Unlike TO-O-CNF surfaces, which have no aldehyde groups either, TO-R-CNF stimulated cells, in osteogenic medium, to have higher alkaline phosphatase activity and to form more biomineralization than TCP and TO-CNF groups. These findings indicate that the presence of aldehyde groups (280 ± 14 μmol/g) on the surface of TEMPO-oxidized CNFs might have little or no effect on attachment, proliferation, and osteogenic differentiation of MSCs.

Quantitative Measurement of Multiprotein Nanoparticle Interactions Using NMR Spectroscopy.

An effective intensity-based reference is a cornerstone for quantitative nuclear magnetic resonance (NMR) studies, as the molecular concentration is encoded in its signal. In theory, NMR is well suited for the measurement of competitive protein adsorption onto nanoparticle (NP) surfaces, but current referencing systems are not optimized for multidimensional experiments. Presented herein is a simple and novel referencing system using 15N tryptophan (Trp) as an external reference for 1H-15N 2D NMR experiments. The referencing system is validated by the determination of the binding capacity of a single protein onto gold NPs. Then, the Trp reference is applied to protein mixtures, and signals from each protein are accurately quantified. All results are consistent with previous studies, but with substantially higher precision, indicating that the Trp reference can accurately calibrate the residue peak intensities and reduce systematic errors. Finally, the proposed Trp reference is used to kinetically monitor in situ and in real time the competitive adsorption of different proteins. As a challenging test case, we successfully apply our approach to a mixture of protein variants differing by only a single residue. Our results show that the binding of one protein will affect the binding of the other, leading to an altered NP corona composition. This work therefore highlights the importance of studying protein-NP interactions in protein mixtures in situ, and the referencing system developed here enables the quantification of binding kinetics and thermodynamics of multiple proteins using various 1H-15N 2D NMR techniques.

Nanotube-decorated hierarchical tantalum scaffold promoted early osseointegration

This study aimed to fabricate a hierarchical tantalum scaffold mimicking natural bone structure to enhance osseointegration. Porous tantalum scaffolds (p-Ta) with microgradients were fabricated by selective laser melting according to a computer-aided design model. Electrochemical anodization produced nanotubes on the p-Ta surface (p-Ta-nt). SEM verified the construction of a unique nanostructure on p-Ta-nt. Contact angle and protein adsorption measurements demonstrated that p-Ta-nt have enhanced hydrophilicity and protein absorption. MC3T3-E1 preosteoblasts showed increased filamentous pseudopods and comparable cell proliferation when cultured on p-Ta-nt. Osteogenic marker gene (Osterix, Runx2, COL-I) transcription was significantly upregulated in MC3T3-E1 cells cultured on p-Ta-nt after 7 days. After implantation into the femurs of New Zealand white rabbits for 2 weeks, histological examination found improved early osseointegration in the p-Ta-nt group. This study showed that a hierarchical tantalum structure could enhance early osteogenic effects in vitro and in vivo.

Dynamics of long-term protein aggregation on low-fouling surfaces

Understanding the mechanisms of protein interactions with solid surfaces is critical to predict how proteins affect the performance of materials in biological environments. Low-fouling and ultra-low fouling surfaces are often evaluated in short-term protein adsorption experiments, where ‘short-term’ is defined as the time required to reach an initial apparent or pseudo-equilibrium, which is usually less than 600 s. However, it has long been recognized that these short-term observations fail to predict protein adsorption behavior in the long-term, characterized by irreversible accumulation of protein on the surface. This important long-term behavior is frequently ignored or attributed to slow changes in surface chemistry over time—such as oxidation—often with little or no experimental evidence. Here, we report experiments measuring protein adsorption on “low-fouling” and “ultralow-fouling” surfaces using single-molecule localization microscopy to directly probe protein adsorption and desorption. The experiments detect protein adsorption for thousands of seconds, enabling direct observation of both short-term (reversible adsorption) and long-term (irreversible adsorption leading to accumulation) protein-surface interactions. By bridging the gap between these two time scales in a single experiment, this work enables us to develop a single mathematical model that predicts behavior in both temporal regimes. The experimental data in combination with the resulting model provide several important insights: (1) short-term measurements of protein adsorption using ensemble-averaging methods may not be sufficient for designing antifouling materials; (2) all investigated surfaces eventually foul when in long-term contact with protein solutions; (3) fouling can occur through surface-induced oligomerization of proteins which may be a distinct step from irreversible adsorption; and (4) surfaces can be designed to reduce oligomerization or the adsorption of oligomers, to prevent or delay fouling.

Facile method for covalent-bonding coating of crosslinked silicone layer onto poly(ester-urethane) surface to improve tensile properties and hemocompatibility

In this paper, a facile method was provided to prepare covalent-bonding coating of crosslinked silicone (CLS) layer onto the poly(ester-urethane) (PEU) surface for improving its properties. The PEU containing well-ordered hard segments was prepared by the direct chain extension of poly(ε-caprolactone) with diurethane diisocyanate (HBH), and the PEU films were acquired via solvent evaporation. The CLS layer was covalently coated onto the film surface by a three-step chemical treatment: the -NCO groups were first introduced onto the PEU film surface by chemical treatment of the film with hexamethylene diisocyanate; the free NCO groups were subsequently coupled with the NH2 groups of γ-aminopropyltriethoxysilane to fix the active alkoxysilane groups on the surface; finally, the alkoxysilane groups self-crosslinked to produce a CLS coating on the PEU surface (PEU-Si). FT-IR, XPS and SEM were used to characterize the modified surface. The studies on physicochemical properties of the films manifested that the PEU-Si film exhibited higher thermal stability, excellent tensile properties (broken elongation: 1389 %; ultimate strength: 41.6 MPa), and slower in vitro hydrolytic degradation (only 16.0 % weight loss after 18 months of degradation) than PEU film. The hemocompatibility evaluation of film surface was performed by protein adsorption and platelet adhesion measurements, and the results showed that the CLS-coated surface strongly inhibited protein adsorption (2.92 μg/cm2) and platelet adhesion (2,129 platelets/mm2). The PEU-Si exhibited excellent tensile properties, slow degradability, and superior surface hemocompatibility, indicating promise as a long-term implant.

Controlled structure and hydrophilic property of polymethylhydrosiloxane thin films attached on silicon support and modified with phosphorylcholine group

Thin films of crosslinked polymethylhydrosiloxane (PMHS) are grafted on silica using sol-gel process allowing subsequent coupling of molecules by hydrosilylation grafting to Si–H groups. The aim was to fabricate stable polysiloxane film on solid supports with well-characterized functionalities and surface properties for various applications. The pristine PMHS surface was modified with phosphorylcholine group by reaction at room temperature or 40 °C catalyzed by Karstedt's catalyst with a phospholipid molecule containing linoleoyl chains to form hydrophilic surfaces as a model. The purpose was to characterize precisely the structure and composition of virgin and treated PMHS films in relation with the wetting properties by using IR and X-ray photoelectron spectroscopies (XPS). The surface properties were studied by pure water contact angle measurements in air by using sessile drops and captive bubbles, atomic force microscopy, and scanning electron microscopy. The modified surfaces are highly hydrophilic with a contact angle varying from 40 to 0°. Analysis of the composition of the surface showed that the reaction of linoleoyl chains gives the 2:1 (Si:phospholipid) hydrosilylation adducts resultant from one fold addition of methylhydrosilane on each linoleoyl chain. The graft yield in terms of molecular percentage determined by XPS in various conditions was correlated in good agreement with wetting properties and previous protein adsorption measurements for comparison purpose.

Resistance to Long-Term Bacterial Biofilm Formation Based on Hydrolysis-Induced Zwitterion Material with Biodegradable and Self-Healing Properties.

Long-term resistance of biomaterials to the bacterial biofilm formation without antibiotic or biocide is highly demanded for biomedical applications. In this work, a novel biodegradable biomaterial with excellent capability to prevent long-term bacterial biofilm formation is prepared by the following two steps. Ethylcarboxybetaine ester analogue methacrylate (ECBEMA), poly(ethylene glycol) monomethacrylate (PEGMA), and 3-methacryloxypropyletris(trimethylsiloxy)silane (TRIS) were copolymerized to obtain p(ECBEMA-PEGMA-TRIS) (PEPT). Then, PEPT was cross-linked by isocyanate-terminated polylactic acid (IPDI-PLA-IPDI) to obtain the final PEPTx-PLAy (x and y are the number-average molecular weights (Mn) of PEPT and PLA, respectively) with optimal mechanical strength and adjustable surface regeneration rate. Static contact angle measurement, protein adsorption measurement, and attenuated total reflectance infrared (ATR-IR) results show that the PEPT19800-PLA800 film surface can generate a zwitterionic layer to resist nonspecific protein adsorption after surface hydrolysis. Quartz crystal microbalance with dissipation (QCM-D) results indicates that the PEPT19800-PLA800 film can undergo gradual degradation of the surface layer at the lowest swelling rate. Particularly, this material can efficiently resist the bacterial biofilm formation of both Gram-positive bacteria and Gram-negative bacteria over 14 and 6 days, respectively. Moreover, the material also shows an ideal self-healing feature to adapt to harsh conditions. Thus, this nonfouling material shows great potential in biomedical applications and marine antifouling coatings without antibiotic or biocide.

Anti-biofouling of 2-acrylamido-2-methylpropane sulfonic acid grafted cellulose acetate membranes used for water desalination

Cellulose acetate based membranes are used for water desalination. Cellulose diacetate (CDA) was prepared from cellulose powder. Reverse osmosis (RO) membranes were prepared on polyester sheets using CDA based through phase inversion technique. The structural, morphological and hydrophilic properties of the prepared membranes were characterized by Fourier transform infrared spectroscopy (FTIR), Nuclear magnetic resonance (NMR), scanning electron microscopy (SEM), and contact angle measurements. The FTIR and NMR revealed the presence of the carbonyl groups and degree of substitution (DS) in CDA. Modified membranes by Grafted to improve the performance and the anti-biofouling properties of cellulose acetate reverse osmosis (RO) membranes. The anti-biofouling properties were studied by measurements of static protein adsorption. The effect of grafting the membrane on the salt rejection and water flux was studied using a cross flow RO unit. The results indicated that 15 wt% of 2-acrylamido-2-methylpropanesulfonic acid (AMPS)Grafted membranes have lower adsorption of protein and microbes, in addition to increased salt rejection to 99.24 % and water flux to 17.12 l/m2 h.

Interaction of endothelial cells with plasma-polymer modified surfaces

Endothelialisation of implantable vascular grafts and stents is directed by the adsorbed protein layer. Chemical and mechanical cues sensed at the biomaterial surface by endothelial cells determine their attachment, survival and proliferation. Given the multiplicity of possible interactions we describe the novel application of live cell imaging, factorial analysis and single molecule imaging to investigate higher order interactions between surface chemistry and adsorbed proteins influencing endothelial cell (EC) adhesion dynamics. EC fates were tracked by time-lapse imaging of cell contact area on plasma polymer modified surfaces. The combinatorial effect of plasma polymer chemistry and adsorbed proteins was characterised by factorial experimental design and analysis. Single molecule imaging and counting was used for the first time to quantify binding of fluorescently labelled albumin to plasma polymerized surfaces: Modified glass surfaces with thin plasma polymer coatings rich in primary amine groups had a high affinity for albumin (~2, 650 molecules/μm2) while plasma polymer surfaces functionalised with hydroxyl groups had very low levels of albumin binding (22–30 molecules/μm2). Plasma polymer coatings rich in primary amine groups also promoted endothelial cell adhesion and was superior to tissue culture plastic. An interaction between albumin, heparin and fibronectin promoted adhesion to amine plasma polymer coated glass, while hydroxyl plasma polymer coated glass prevented EC attachment. Tracking cell-specific interactions with the adsorbed protein layer by time-lapse microscopy is more predictive of in vivo cellular responses to biomaterials compared to studies that only measure protein adsorption.

Sample Preparation in Quartz Crystal Microbalance Measurements of Protein Adsorption and Polymer Mechanics.

In this study, we present various examples of how thin film preparation for quartz crystal microbalance experiments informs the appropriate modeling of the data and determines which properties of the film can be quantified. The quartz crystal microbalance offers a uniquely sensitive platform for measuring fine changes in mass and/or mechanical properties of an applied film by observing the changes in mechanical resonance of a quartz crystal oscillating at high frequency. The advantages of this approach include its experimental versatility, ability to study changes in properties over a wide range of experimental time lengths, and the use of small sample sizes. We demonstrate that, based on the thickness and shear modulus of the layer deposited on the sensor, we can acquire different information from the material. Here, this concept is specifically exploited to display experimental parameters resulting in mass and viscoelastic calculations of adsorbed collagen on gold and polyelectrolyte complexes during swelling as a function of salt concentration.

Sample Preparation in Quartz Crystal Microbalance Measurements of Protein Adsorption and Polymer Mechanics

Sample Preparation in Quartz Crystal Microbalance Measurements of Protein Adsorption and Polymer Mechanics

Protein adsorption measurements on low fouling and ultralow fouling surfaces: A critical comparison of surface characterization techniques

Ultralow protein fouling behavior is a common target for new high-performance materials. Ultralow fouling is often defined based on the amount of irreversibly adsorbed protein (< 5 ng cm−2) measured by a surface ensemble averaging method. However, protein adsorption at solid interfaces is a dynamic process involving multiple steps, which may include adsorption, desorption, and irreversible protein denaturation. In order to better optimize the performance of antifouling surfaces, it is imperative to fully understand how proteins interact with surfaces, including kinetics of adsorption and desorption, conformation, stability, and amount of adsorbed proteins. Defining ultralow fouling surfaces based on a measurement at or near the limit of detection of a surface-averaged measurement may not capture all of this behavior. Single-molecule microscopy techniques can resolve individual protein-surface interactions with high temporal and spatial resolution. This information can be used to tune the properties of surfaces to better resist protein adsorption. In this work, we demonstrate how combining surface plasmon resonance, X-ray photoelectron spectroscopy, atomic force microscopy, and single-molecule localization microscopy provides a more complete picture of protein adsorption on low fouling and ultralow fouling polyelectrolyte multilayer and polymer brush surfaces, over different regimes of protein concentration. In this case, comparing the surfaces using surface plasmon resonance alone is insufficient to rank their resistance to protein adsorption. Our results suggest a revision of the accepted definition of ultralow fouling surfaces is timely: with the advent of time-resolved studies of protein adsorption kinetics at the single-molecule level, it is neither necessary nor sufficient to rely on a surface averaging techniques to qualify ultralow fouling surfaces. Since protein adsorption is a dynamic process, understanding how surface properties affect the kinetics of protein adsorption will enable the design of future generations of advanced antifouling materials. Statement of significanceThe design of ultralow fouling surfaces is often optimized based on a single surface-averaging technique measuring the amount of irreversibly adsorbed protein. This work provides a critical comparison of alternative techniques for evaluating protein adsorption on low fouling and ultralow fouling surfaces, and demonstrates how additional information about the dynamics of protein-surface interactions at the interface can be obtained by application of single-molecule microscopy. This approach could be used to better elucidate mechanisms of protein resistance and design principles for advanced ultralow fouling materials.

Fibrinogen and albumin adsorption profiles on Ni-free Zr-based thin film metallic glass

In this study, we assessed the fibrinogen and albumin adsorption performance of Ni-free Zr-based (Zr53Cu33Al9Ta5) thin-film metallic glass (TFMG). Quartz crystal microbalance (QCM) system and fluorescence spectrophotometer were used to obtain protein adsorption measurements, using two different substrates (QCM and glass). Test samples were fabricated by coating both of the substrates with TFMG and Ti (widely used in implants) for comparison. In dynamic QCM as well as static fluorescence spectrophotometer tests, the TFMG achieved a lower fibrinogen to albumin adsorption ratio. The TFMG coating also exhibited high hydrophobicity (water contact angle of 96°) with low surface roughness of 1.06 nm and surface free energy of 35.3 mN/m. This is a clear demonstration of the excellent hemocompatibility of Ni-free Zr-based TFMG.

Commercial polysulfone membranes pretreated with ethanol and NaOH: Effects on permeability, selectivity and antifouling properties

This study explores the effect of three simple pretreatment methods – hot water, ethanol (EtOH) and sodium hydroxide (NaOH) – on the performance of commercial polysulfone (PSf) membranes. 100% EtOH and 1 M NaOH pretreated membranes exhibited increases of, respectively, 600% and 50% of water permeability compared to the hot water pretreated membrane (control). Such increases were partially ascribed to the removal of preservatives i.e. glycerol and polyvinylpyrrolidone (PVP) in the chemically pretreated membranes, as confirmed by FTIR spectra. The dramatic increase of water permeability in the ethanol pretreated membranes was also explained by pore enlargement, which was confirmed by decreased retention of dextran of selected molecular weights. Protein (BSA and pepsin) filtration experiments confirmed the occurrence of higher flux during filtration when using ethanol and NaOH pretreated membranes, though fouling caused during filtration was more severe and more difficult to remove in ethanol pretreated membranes compared to the control. Static protein adsorption measurements confirmed that ethanol pretreated membranes had an enhanced propensity to adsorb more protein. Water permeability tests performed after maintaining the pretreated membranes in water for several days confirmed that the changes induced by the selected pretreatments persist after at least 7 days. In general these results show that the studied pretreatment methods can be used to tailor the properties of commercial polysulfone membranes in terms of permeability and retention to a certain extent and in a simple, inexpensive manner.

Surface-functionalization of isotactic polypropylene via dip-coating with a methacrylate-based terpolymer containing perfluoroalkyl groups and poly(ethylene glycol)

Isotactic polypropylene (PP) is one of the most popular plastics. However, the remarkably low surface energy of PP prevents the surface functionalization of PP. We studied the surface functionalization of PP by dip-coating with a maleic anhydride-grafted chlorinated polypropylene (MPO)/methacrylate-based terpolymer mixture. A methacrylate-based terpolymer (PMFP) was synthesized, which contained perfluoroalkyl (Rf)-conjugated monomers and poly(ethylene glycol)-conjugated monomers. Tape-peeling tests revealed that MPO successfully immobilized PMFP on a PP surface, although PMFP was less adhesive to PP. X-ray photoelectron spectroscopy (XPS), contact angle, and protein adsorption measurements revealed that the Rf groups and PEG chains in PMFP were segregated to the outermost surface of the dip-coated layer. The surface segregation of these moieties produced a low-fouling surface on the PP substrate. In addition, we synthesized a terpolymer that contained Rf groups and PEG chains with carboxy groups at their termini (PMFB) and used it to dip-coat a PP substrate. The surface segregation of side chains in PMFB induced the presentation of carboxy groups at the outermost surface, which were used as reactive sites for enzyme immobilization. We studied the surface functionalization of PP by dip-coating with a maleic anhydride-grafted chlorinated polypropylene (MPO) /methacrylate-based terpolymer mixture. A methacrylate-based terpolymer (PMFP) was synthesized, which contained perfluoroalkyl (Rf)-conjugated monomers and poly(ethylene glycol)-conjugated monomers. We found that the presence of MPO aided the adhesion of the terpolymer to the PP surface and that the coated PP surface exhibited low protein-fouling properties. In addition, the dip-coating of a terpolymer containing reactive groups (–COOH) with MPO caused the reactive groups to be presented at an outermost surface.

Effects of calcium and phosphorus incorporation on the properties and bioactivity of TiO2 nanotubes.

In this study, we investigate the formation of calcium and phosphorus-doped TiO2 nanotubes, produced by potentiostatic anodization of Ti in viscous electrolyte-containing HF and Ca/P ions. Characterization of the produced oxide layer was conducted using scanning electron microscopy, glancing-angle X-ray diffraction, X-ray photoelectron spectroscopy, contact angle, and protein adsorption measurements. Adipose-derived stem cells were used to study material cytotoxicity, cell viability and proliferation, and cell morphology and growth. To evaluate the adipose-derived stem-cell differentiation, we investigated the expression of osteocalcin and osteopontin by cells as well as calcium mineralization. Results show that it was possible to produce a superhydrophilic titanium oxide nanotube layer with incorporation of Ca and P ions. The presence of Ca and P in the oxide layer not only improved the cell adhesion and proliferation but also stimulated the production of key marker proteins indicating differentiation of cells.

Resisting Protein but Promoting Cell Adhesion by Choline Phosphate: A Comparative Study with Phosphorylcholine

Zwitterionic materials are now widely used to fabricate various functionalized surfaces for biomedical applications due to their excellent non-fouling properties. As a newly-discovered zwitterionic material, choline phosphate (CP) was reported to be cell-adhesive, which makes it different from traditional non-fouling zwitterionic materials such as sulfobetaine, carboxybetaine and phosphorylcholine (PC). However, there are still few reports about choline phosphate, and to the best of our knowledge, a direct comparative study between choline phosphate and conventional zwitterionic materials has not been reported to date. To further investigate the properties of choline phosphate, a comparative study was conducted and the widely-reported zwitterionic phosphorylcholine, which has the same chemical component but opposite charge orientation with choline phosphate, was employed as a control. For this purpose, CP and PC-functionalized surfaces were prepared by surface-initiated atom transfer radical polymerization (Si-ATRP), and their non-fouling properties were probed by protein adsorption and eukaryotic cell adhesion measurement. The result shows that CP-functionalized surfaces exhibit almost equivalent amounts of adsorbed proteins to that of PC, but they are more beneficial to cells initial adhesion and further spreading in serum-free medium, indicating that CP has a promising prospect of application in tissue engineering.